Biology subjects

Brown, J. S.

Publications and source records attributed to Brown, J. S..

At least 19 recordsLinked to original sources

bioRxiv2026

Wasting is essential: cachexia is driven by a wound-healing adaptation

Background: Several explanations have been given for why cachexia occurs in the presence of severe acute wounds and chronic diseases. None of these explanations address the scope of clinical settings in which cachexia has been observed, nor do they account for the failure of nutritional support to reverse cachexia. We hypothesize that induced muscle wasting (IMW) is an adaptation to support wound healing, and when a wound does not heal, prolonged IMW leads to cachexia. Methods: To evaluate this hypothesis, we developed a mathematical model of IMW. In this model, we compared the time needed for wound healing when IMW was present and when it was inhibited. We also measured the relationship between the pathogen growth rate and final patient muscle mass for chronic diseases such as cancer and tuberculosis, demonstrating the onset of cachexia. Finally, we simulated the effects of nutritional support and contrasted the outcomes of when IMW was present and when it was inhibited. Results: Acute wound healing was ~2-6 times longer without IMW than with IMW. For chronic wounds, steady-state muscle mass strongly depended on the growth rate of the pathogenic agent. Inhibiting IMW was more effective at preventing acute muscle mass loss than nutritional support, and the benefit of nutritional support was enhanced by inhibiting IMW. Nutritional support also increased the speed of healing acute wounds when IMW was inhibited. For cancer with high growth rate, giving nutritional support alone had a negligible effect on steady-state muscle mass. But when IMW was inhibited, muscle mass loss was attenuated. With IMW inhibition, nutritional support with the normal diet was able to abrogate muscle mass loss entirely. Conclusions: Inhibiting IMW slows the healing of acute wounds, supporting our hypothesis that IMW is an adaptation to support rapid wound healing. Our experiments demonstrate that nutritional support is insufficient to prevent severe muscle mass loss while IMW continues. But when IMW is inhibited, nutritional support restores normal levels of muscle mass for both acute and chronic wounds, as well as increasing the rate of wound healing. Discovering and developing methods to inhibit IMW for clinical use will be essential to restore patient muscle mass, ease suffering, and reduce mortality.

systems biology↗

bioRxiv2026

Initial tumor composition shapes resistance evolution and treatment outcomes in non-small cell lung cancer

Drug resistance is a leading cause of treatment failure in non-small cell lung cancer (NSCLC), yet how resistance evolves during treatment and whether its fitness consequences depend on tumor composition remains poorly understood. Using a game-theoretic mathematical model fitted to longitudinal in-vitro data from alectinib-sensitive and alectinib-resistant H3122 NSCLC cells grown under different treatment and microenvironmental conditions, we found that the fitness effect of evolving resistance depended critically on the initial proportion of resistant cells in the tumor. When resistant cells were initially rare, resistance evolved faster and increasing resistance was associated with a growth advantage. When resistant cells were initially frequent, increasing resistance was associated with a fitness cost. In both cases, increasing resistance eroded treatment efficacy. In the gain-of-resistance regime, stabilization therapy could maintain a stable tumor equilibrium only if resistant cells were excluded. Maximum tolerated dosing was not always optimal for maximizing time to progression; intermediate doses performed better when they kept the initial tumor growth rate close to zero. These results suggest that evolutionary therapy for NSCLC should account not only for the abundance of resistant cells, but also for how resistance is evolving and what fitness consequences it currently carries in individual patients.

cancer biology↗

bioRxiv2026

Accounting for DNA Recovery and Cell Culturability Enhances Quantitative Compatibility of Molecular and Legiolert Assays for Legionella pneumophila

Disagreement between molecular and culture-based assays for Legionella pneumophila detection is widely reported, yet comparisons have largely been based on direct assay-derived concentrations or binary positive/negative outcomes. However, it remains unclear whether molecular-culture disagreement reflects concentration-level incompatibility or unaccounted methodological and physiological differences related to DNA recovery and cell culturability. In this study, we also observed disagreement between molecular and Legiolert assays in source and finished drinking water samples collected from eight full-scale drinking water systems across the United States. Molecular thresholds adjusted for DNA recovery and cell culturability only partially resolved these discrepancies. We therefore developed a probabilistic Monte Carlo framework that incorporates sample-specific DNA recovery and cell culturability to evaluate the quantitative consistency of culturable L. pneumophila concentrations estimated by molecular and Legiolert assays. Quantitatively consistent and inconsistent samples occurred across both binary concordant and discordant classifications, demonstrating that positive/negative agreement poorly reflects concentration-level comparability. Overall, molecular and Legiolert assays showed strong quantitative consistency once sample-specific DNA recovery and cell culturability were considered. A small proportion of persistent inconsistencies at specific sampling sites, coupled with atypical microbial indicators, suggest that sample heterogeneity likely contributed to the remaining discrepancies. These findings demonstrate that integrating DNA recovery and cell culturability enhanced quantitative consistency between molecular and Legiolert assays and supports the use of molecular methods as rapid quantitative tools to complement culture-based L. pneumophila monitoring. SynopsisAccounting for DNA recovery and cell culturability revealed broad quantitative compatibility between molecular and Legiolert assays for Legionella pneumophila in drinking water. TOC O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=88 SRC="FIGDIR/small/739452v2_ufig1.gif" ALT="Figure 1"> View larger version (26K): org.highwire.dtl.DTLVardef@1e01cdcorg.highwire.dtl.DTLVardef@86bbc2org.highwire.dtl.DTLVardef@190fc66org.highwire.dtl.DTLVardef@1aa9ec8_HPS_FORMAT_FIGEXP M_FIG C_FIG

microbiology↗

bioRxiv2026

Cancer Evolvability Determines Therapy Outcomes

Cancer progression following treatment failure is an evolutionary process in which therapy acts as a selection pressure driving Darwinian selection on heritable variation to favor resistant clones. This ability to generate variation, i.e., the cancers evolvability, is a key determinant of how rapidly tumors adapt to therapy. Here, we present an evolutionary game-theoretic model to evaluate how evolvability shapes resistance dynamics under two treatment modalities: targeted therapy and chemotherapy. We first compare cancer populations with fixed evolvabilities: low or high. Targeted therapy imposes a steep selection gradient, enabling rapid resistance evolution, while chemotherapy exerts a flatter gradient but drives tumors toward more extreme resistance strategies. We show that targeted therapy works better in low-evolvability cancers, whereas chemotherapy better controls high-evolvability populations. We then extend the model to incorporate facultative evolvability in which cancer cells dynamically adjust their evolvability in response to therapy-induced stress in which cells fine-tune the trade-off between acquiring higher resistance and limiting the costs of resistance and evolvability. The latter strategy sustains a higher tumor burden than fixed-evolvability populations. To address the challenges of facultative evolvability for therapy efficacy, we develop and simulate an evolutionary double bind using sequential cycles of chemotherapy and targeted therapy. With an appropriate sequence and timing, this strategy can drive cancer cells with facultative evolvability to extinction. Our results highlight the importance of evolvability in shaping treatment response and underscore the need to incorporate evolutionary principles into therapy design.

cancer biology↗

bioRxiv2026

Modeling VEGF and GLUT1 Expression as Coadapted Foraging Strategies in Cancer

Natural selection acting on cancer cells within their tumor microenvironment should favor cells with fast or efficient nutrient uptake strategies. Here, we develop and analyze a game-theoretic model focusing on the coadaptation between two foraging traits: vascular endothelial growth factor (VEGF) and glucose transporter 1 (GLUT1). Studies show that VEGF and GLUT1 are often co-expressed and are associated with more aggressive tumor phenotypes and poor clinical prognosis. VEGF is a diffusible paracrine factor that recruits blood vessels towards neighborhoods of cancer cells (angiogenesis). GLUT1 is a cell-surface transporter that enables the uptake of nutrients, especially glucose. We model these strategies operating at different scales: VEGF influences resource availability at the neighborhood level, while GLUT1 determines resource uptake at the cellular level. For VEGF, we introduce a resource-sharing continuum. With no resource sharing, cells access resources in proportion to their VEGF contribution. With uniform sharing, cells have equal access to resources, regardless of their VEGF contribution. The former leads to a tragedy of the commons and overproduction of VEGF. The latter yields a public goods game with moderate VEGF expression matching a group optimum. GLUT1 expression mediates uptake of resources recruited by VEGF and is largely independent of the degree of resource sharing. Therapeutically, both VEGF and GLUT1 inhibitors are more effective in high resource-sharing neighborhoods and less so as resource sharing declines. Overall, inhibition of GLUT1-mediated uptake emerges as more effective. The model, perhaps the first to consider VEGF and GLUT1 as coadaptations, emphasizes the need to consider cancer cell traits jointly.

cancer biology↗

bioRxiv2025

Streptococcus pneumoniae accessory capsular genes modulate fitness, pathogenicity and immune evasion

Globally, Streptococcus pneumoniae disproportionally affects children in resource-poor settings, older adults and people living with HIV. Frequently found as an asymptomatic colonizer of the nasopharynx, this versatile pathogen is a prominent cause of pneumonia, meningitis, bacteraemia and otitis media. Recently, a serotype 3 capsule variant (GPSC10-ST700) has expanded in Malawi with enhanced vaccine escape potential. Here, using a mutational and complementation approach, we show that loss of accessory capsular genes in GPSC10-ST700 contribute to increased opsonophagocytic resistance in this lineage. Although originally thought to be nonfunctional pseudogenes, we show that these genes modulate fitness and the global phosphoproteome in serotype 3 strains. These findings highlight that vaccine escape may be mediated through variations in the pneumococcal capsular locus that enhance fitness, pathogenicity and immune evasion, without capsule switching. IMPORTANCEPneumococcal polysaccharide-conjugate vaccines (PCV) target the polysaccharide capsule (CPS), which is a dominant virulence factor. However, current PCVs induce suboptimal protection against serotype 3 strains, which produce a thicker capsule that when released from the bacterial surface, interferes with antibody-mediated bacterial killing and protection. We recently described the clonal expansion of a sequence type (ST) 700-GPSC10 serotype 3 lineage in Malawi post-PCV13 introduction. This lineage is characterized by the absence of at least 6 genes in its cps locus and a distinct antimicrobial resistance (AMR) profile compared to other serotype 3 strains. Here we uncovered a functional role for the accessory capsular genes (acl) in serotype 3, previously considered to be pseudogenes, which modulate capsule production, shedding, serum tolerance, and bacterial fitness. By linking genotype to phenotype, our work provides new insights into the molecular basis of serotype 3 immune evasion, informing the design of more effective pneumococcal vaccines.

microbiology↗

bioRxiv2025

RNA Helicase A promotes small RNA biogenesis and sorting in germ cells

In germ cells, small RNAs function as a defense system to silence invading RNAs like viruses and transposons to protect genome integrity. The ability of small RNAs to robustly silence diverse RNA sequences prompts the question of how endogenous mRNAs avoid this silencing. In C. elegans, small RNAs bound by the Argonaute CSR-1 protect endogenous mRNAs from silencing, while also fine-tuning a subset of these mRNAs. Here, we identify RNA Helicase A (RHA-1) as a key regulator of CSR-1 small RNA biogenesis and function in mRNA fine-tuning. RHA-1 localizes to germ granules dependent on EGO-1, which synthesizes CSR-1 small RNAs. We find RHA-1 promotes small RNA production from the 5 regions of mRNAs and small RNA sorting to CSR-1. Loss of RHA-1 leads to elevated CSR-1 target mRNA levels and compromised fertility. Our study highlights the importance of small RNA regulation, mediated by RHA-1, to protect endogenous gene expression programs and germ cell function.

genetics↗

bioRxiv2025

Analysis of crosslinking sites suggests C. elegnas PIWI Argonaute exhibits flexible conformations for target recognition

Small RNAs play critical roles in gene regulation in diverse processes across organisms. Crosslinking, ligation, and analyses of sequence hybrid (CLASH) experiments have shown PIWI and Argonaute proteins bind to diverse mRNA targets, raising questions about their functional relevance and the degree of flexibility in target recognition. As crosslinking-induced mutations (CIMs) provides nucleotide-resolution of RNA binding sites, we developed MUTACLASH to systematically analyze CIMs in piRNA and miRNA CLASH data in C. elegans. We found CIMs are enriched at the nucleotide positions of mRNA corresponding to the center of targeting piRNAs and miRNAs. Notably, CIMs are also enriched at nucleotides with local pairing mismatches to piRNA. In addition, distinct patterns of CIMs are observed between canonical and non-canonical base pairing interactions, suggesting that the worm PIWI Argonaute PRG-1 adopts distinct conformations for canonical vs. non-canonical interactions. Critically, non-canonical miRNA or piRNA binding sites with CIMs exhibit more regulatory effects than those without CIMs, demonstrating CIM analysis as a valuable approach in assessing functional significance of small RNA targeting sites in CLASH data. Together, our analyses reveal the landscapes of Argonaute crosslinking sites on mRNAs and highlight MUTACLASH as an advanced tool in analyzing CLASH data.

molecular biology↗

bioRxiv2024

Goliath clades and in vivo tracking of clonal dynamics show three phases of UV-induced skin carcinogenesis

While the genetic paradigm of cancer etiology has proven powerful, it remains incomplete as evidenced by the widening spectrum of non-cancer cell-autonomous "hallmarks" of cancer. Studies have demonstrated the commonplace presence of high oncogenic mutational burdens in homeostatically-stable epithelia. Hence, the presence of driver mutations alone does not result in cancer. Here, we reveal additional forces governing the eco-evolutionary dynamics of carcinogenesis. Using a UV-driven mouse model of cutaneous squamous cell carcinoma, we tested our central hypothesis that cancer initiation occurs in three phases: 1) tissue disruption and the emergence of unusually large "goliath" clades (ecological driver), 2) clonal selection within a subset of these goliaths as evidenced by the presence of areas of unusually high local densities of cells (termed "micro-lumps") with higher mutational burdens (evolutionary driver), and 3) emergence of macroscopic lesions. We tracked these tissue level ecological and evolutionary drivers of cancer initiation via in-vivo serial imaging and 3-D reconstruction of fluorescently labeled keratinocyte clades, yielding 25,085 clade measurements from 14 mice over 3 months, and 14,525 clades from 4 of these mice over 6 months. While median and mean clade sizes differed little between UV and non-UV exposure (cc. 65,000 {micro}m3), our ecological survey revealed the emergence of large goliath clades (> cc. 4.2x106 {micro}m3), almost exclusively within UV-exposed skin. Goliath clades emerged by month 2, increased dramatically in number by months 3-4, and plateaued between months 5-6. Goliath clades arose as serpentine structures, intercalating between adjacent keratinocytes. Unexpectedly, targeted DNA sequencing revealed mutations with very low variant allele frequencies within clades, but substantial differences among clades, suggesting that positive selection for these mutations is superfluous to the development of goliaths early in carcinogenesis. scRNAseq analysis of both bulk skin and sorted clades revealed epidermal de-differentiation and immune suppression as early events. By month 7, mutational burdens were significantly larger in goliath clades, particularly for those with micro-lumps. Finally, lesions began to emerge between months 6 and 7, only in UV-exposed skin. To confirm that goliath clades are orders of magnitude more likely to spawn lesions, we randomly selected 21 goliath clades at months 6-7, prior to the time of the emergence of detectable lesions, and followed them over time. Remarkably, 2 of these developed into macroscopic lesions. Our adaptation of the Drake equation estimated the probability of this to be <10-6. Taken together, our results support the presence of three phases of cancer initiation, the earliest of which presage the acquisition of driver mutations and also explains why cancers are relatively rare in relation to the degree of somatic mosaicism present in UV-exposed skin.

cancer biology↗

bioRxiv2024

Migrastatic Therapy as a Potential Game-Changer inAdaptive Cancer Treatment

Adaptive therapy, which anticipates and counters the evolution of resistance in cancer cells, has gained significant traction, especially following the success of the Zhang et al.s protocol in treating metastatic castrate-resistant prostate cancer. While several adaptive therapies have now advanced to clinical trials, none currently incorporates migrastatics, i.e. treatments designed to inhibit cancer cell metastasis. In this study, we propose integrating migrastatics into adaptive therapy protocols and evaluate the potential benefits of using a game-theoretic spatial model. Our results demonstrate that combining adaptive therapy with migrastatics effectively delays the onset of metastasis and reduces both the number and size of metastases across most cancer scenarios analyzed. This approach not only extends the time to the first metastasis but also enhances the overall efficacy of adaptive therapies. Our findings suggest a promising new direction for cancer treatment, where adaptive therapy, in combination with migrastatic agents, can target both the evolution of resistance and the metastatic spread of cancer cells.

cancer biology↗

bioRxiv2024

Evolutionary rescue model informs strategies for driving cancer cell populations to extinction

Cancers exhibit a remarkable ability to develop resistance to a range of treatments, often resulting in relapse following first-line therapies and significantly worse outcomes for subsequent treatments. While our understanding of the mechanisms and dynamics of the emergence of resistance during cancer therapy continues to advance, questions remain about how to minimize the probability that resistance will evolve, thereby improving long-term patient outcomes. Here, we present an evolutionary simulation model of a clonal population of cells that can acquire resistance mutations to one or more treatments. We leverage this model to examine the efficacy of a two-strike "extinction therapy" protocol, in which two treatments are applied sequentially to first contract the population to a vulnerable state and then push it to extinction, and compare it to a combination therapy protocol. We investigate how factors such as the timing of the switch between the two strikes, the rate of emergence of resistant mutations, the doses of the applied drugs, the presence of cross-resistance, and whether resistance is a binary or a quantitative trait affect the outcome. Our results show that the timing of switching to the second strike has a marked effect on the likelihood of driving the cancer to extinction, and that extinction therapy outperforms combination therapy when cross-resistance is present. We conduct an in silico trial that reveals when and why a second strike will succeed or fail. Finally, we demonstrate that our conclusions hold whether we model resistance as a binary trait or as a quantitative, multi-locus trait.

evolutionary biology↗

bioRxiv2024

Additive, substitutable, and antagonistic prey responses to feral and native predator combinations

In captive experiments of Negev Desert granivores, we investigated the ways in which combinations of feral mesocarnivores and native predators alter wild prey behavior. We hypothesized that feral mesocarnivores would have a greater impact on prey energy acquisition, reflected in foraging dynamics, than native predators. Allenbys gerbils (Gerbillus andersoni allenbyi) and greater Egyptian gerbils (Gerbillus pyramidum) were used as prey species, with feral dogs (Canis lupus familiaris), feral cats (Felis catus), barn owls (Tyto alba), and horned vipers (Cerastes gasperetti) as predators. Gerbil perceived risk was measured using optimal patch-use theory, with exposures to tethered predators occurring hourly throughout the night. Some nights, two predators were alternated every other hour. We found that human-commensal predators, particularly feral cats, induced stronger foraging than native predators, such as barn owls. Combined predators caused gerbils to decrease foraging only when a higher-risk predator was introduced, as indicated by higher giving-up densities (GUDs) for the dog and cat combination compared to the dog alone, and a nonsignificant increase compared to the cat alone. The impact of feral cats especially appears to outweigh that of native predators. This highlights the conservation challenges to arid environments where feral cats have become ubiquitous. HighlightsO_LIFeral predators, particularly cats and dogs, elicit stronger anti-predator responses in Negev Desert gerbils than native predators, significantly altering foraging behavior and habitat use. C_LIO_LIThe presence of feral predators in the Negev Desert leads to cumulative impacts on prey species, resulting in increased vigilance and reduced resource acquisition, which suggests a disruption of native predator-prey dynamics. C_LIO_LIThe studys findings emphasize the unique conservation challenges posed by human-commensal species in arid ecosystems, where limited refugia increases the predation risk from invasive and feral species. C_LIO_LIThis research demonstrates how predator-prey interactions in the arid communities tested are impacted by both additive predator effects between native predators, and between feral predators, but when combining feral predators with native ones, the effect is substitutable with feral predators disproportionately influencing prey behavior. C_LI Graphical Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=145 SRC="FIGDIR/small/612031v2_ufig1.gif" ALT="Figure 1"> View larger version (80K): org.highwire.dtl.DTLVardef@1d2b45eorg.highwire.dtl.DTLVardef@3f9490org.highwire.dtl.DTLVardef@32a733org.highwire.dtl.DTLVardef@17f2111_HPS_FORMAT_FIGEXP M_FIG C_FIG

animal behavior and cognition↗

bioRxiv2024

The CSF transcriptome in pneumococcal meningitis reveals compartmentalised host inflammatory responses associated with mortality.

AbstractO_ST_ABSBackgroundC_ST_ABSPneumococcal meningitis (PM) has persistently poor clinical outcomes, especially in sub- Saharan Africa. To better characterise the inflammatory response and identify factors associated with mortality we compared paired peripheral blood and cerebrospinal fluid (CSF) transcriptomes before the initiation of antibiotics in Malawian adults with proven PM. ResultsBlood transcriptional profiles were obtained in 28 patients with PM, with simultaneous paired with CSF profiles available for 13 patients. 15/28 (52%) patients died. Comparison of the transcriptome between CSF and blood compartments showed upregulation of 2293 differentially expressed genes in CSF and 909 in blood; enriched pathways in CSF included inflammasome activity and neutrophil migration/activation in the CSF, contrasting with enrichment for pathways including platelet and endothelial activation, cell cycle, cytokine release and oxidative stress in the blood transcriptome. Comparison of CSF profiles between survivors and non-survivors revealed 1829 differentially expressed genes, non- survivor CSF was enriched for multiple innate inflammatory pathways, including IL-17A and Type 1 interferons and proteolysis. In contrast, minimal transcriptomic differences between outcome groups were detected in blood. ConclusionInflammation in PM is characterised by compartmentalised responses in blood and CSF. Poorer outcomes are associated with an dysregulated innate immune host response to S. pneumoniae in the CSF compartment.

immunology↗

bioRxiv2024

The Streptococcus pneumoniae transcriptome in patient cerebrospinal fluid identifies novel virulence factors required for meningitis

To better understand Streptococcus pneumoniae pathogenesis we performed RNA sequencing on cerebrospinal fluid (CSF) from meningitis patients to identify bacterial genes expressed during invasion of the central nervous system. Comparison to transcriptome data for serotype 1 S. pneumoniae cultured in ex vivo human CSF defined a subset of 57 genes with high expression during human meningitis. Deletion of two of the most highly expressed genetic loci, bgaA (encodes for a {beta}-galactosidase) or the SP_1801-5 putative stress response operon, resulted in S. pneumoniae strains still able to transmigrate the blood brain barrier but which were more susceptible to complement opsonisation and unable to maintain brain infection in a murine meningitis model. In 1144 meningitis patients, infection with bgaA containing S. pneumoniae strains was associated with a higher mortality (22% versus 14% p=0.02). These data demonstrate that direct bacterial RNAseq from CSF can identify previously undescribed S. pneumoniae virulence factors required for meningitis pathogenesis.

microbiology↗

bioRxiv2024

Evolutionary double-bind treatment using radiotherapy and NK cell-based immunotherapy in prostate cancer

Evolution-informed therapies exploit ecological and evolutionary consequences of drug resistance to inhibit the expansion of treatment-resistant populations and prolong time to progression. One strategy, termed an evolutionary double-bind, uses an initial therapy to elicit a specific adaptive response by the cancer cells, which is then selectively targeted by a follow-on therapy. Here we examine the combination of radiation therapy and immunotherapy as a quantifiable double-bind strategy. Radiotherapy (RT) induces lethal double-strand DNA breaks, but cancer cells can adapt by upregulating DNA damage response pathways. While this evolutionary strategy increases resistance to DNA damaging agents, it also results in enhanced expression of natural killer (NK) cell ligands potentially increasing vulnerability to an immune response. Using a radiation-resistant human prostate carcinoma cell line (22Rv1), we demonstrate that RT-resistant cells upregulate NK cell ligands, including major histocompatibility complex class I chain-related protein A/B (MICA/B), and poliovirus receptors (PVR1, PVRL2) with a 2-fold increase in sensitivity to NK cell mediated killing. We investigated this potential evolutionary double bind through in vitro studies and evolution-based mathematical models. Radiotherapy alone slowed overall growth but strongly selected for RT-resistant cells. NK cell therapy alone suppressed the RT-resistant population but with a surviving population of radiation-sensitive cells. These dynamics were framed mathematically, and model simulation predicted optimal tumour control would be achieved through initial RT rapidly followed by NK-based immunotherapy. Subsequent experiments confirmed the model prediction. We conclude that radiotherapy and NK cell-based immunotherapy produces an evolutionary double bind that can be exploited in heterogenous tumours to limit RT resistance.

cancer biology↗

bioRxiv2023

Cell Types or Cell States? An Investigation of Adrenergic and Mesenchymal Cell Phenotypes in Neuroblastoma

Neuroblastoma is a pediatric cancer that exhibits two cellular phenotypes: adrenergic (ADRN) and mesenchymal (MES). ADRN is differentiated and therapy-sensitive, while MES is less differentiated with elevated therapy resistance. To understand neuroblastoma and its treatment response, it is important to elucidate how these phenotypes impact the eco-evolutionary dynamics of cancer cell populations and whether they represent distinct cell types or dynamic cell states. Here, we show that neuroblastoma cells undergo an ADRN to a MES phenotypic switch under chemotherapy treatment. We use a strong inference approach to generate four hypotheses on how this switch may occur: cell types without resistance, cell types with resistance, cell states without resistance, and cell states with resistance. For each of these hypotheses, we create theoretical models to make qualitative predictions about their resulting eco-evolutionary dynamics. Our results provide a framework to further experimentally determine whether ADRN and MES phenotypes are distinct cell types or dynamic cell states.

cancer biology↗

bioRxiv2023

Bacterial surface lipoproteins mediate epithelial microinvasion by Streptococcus pneumoniae

Streptococcus pneumoniae, a common coloniser of the upper respiratory tract, invades nasopharyngeal epithelial cells without causing disease in healthy people. We hypothesised that surface expression of pneumococcal lipoproteins, recognised by the innate immune receptor TLR2, mediate epithelial microinvasion. Mutation of lgt in serotype 4 (TIGR4) and serotype 6B (BHN418) pneumococcal strains abolishes the ability of the mutants to activate TLR2 signalling. Loss of lgt also led to concomitant decrease in interferon signalling triggered by the bacterium. However, only BHN418 lgt::cm but not TIGR4 lgt::cm was significantly attenuated in epithelial adherence and microinvasion compared to their respective wild-type strains. To test the hypothesis that differential lipoprotein repertoires in TIGR4 and BHN418 lead to the intraspecies variation in epithelial microinvasion, we employed a motif-based genome analysis and identified an additional 525 a.a. lipoprotein (pneumococcal accessory lipoprotein A; palA) encoded by BHN418 that is absent in TIGR4. The gene encoding palA sits within a putative genetic island present in [~]10% of global pneumococcal isolates. While palA was enriched in carriage and otitis media pneumococcal strains, neither mutation nor overexpression of the gene encoding this lipoprotein significantly changed microinvasion patterns. In conclusion, mutation of lgt attenuates epithelial inflammatory responses during pneumococcal-epithelial interactions, with intraspecies variation in the effect on microinvasion. Differential lipoprotein repertoires encoded by the different strains do not explain these differences in microinvasion. Rather, we postulate that post-translational modifications of lipoproteins may account for the differences in microinvasion. IMPORTANCEStreptococcus pneumoniae (pneumococcus) is an important mucosal pathogen, estimated to cause over 500,000 deaths annually. Nasopharyngeal colonisation is considered a necessary prerequisite for disease, yet many people are transiently and asymptomatically colonised by pneumococci without becoming unwell. It is therefore important to better understand how the colonisation process is controlled at the epithelial surface. Controlled human infection studies revealed the presence of pneumococci within the epithelium of healthy volunteers (microinvasion). In this study, we focused on the regulation of epithelial microinvasion by pneumococcal lipoproteins. We found that pneumococcal lipoproteins induce epithelial inflammation but that differing lipoprotein repertoires do not significantly impact the magnitude of microinvasion. Our results highlight the potential importance of the post-translational modification of lipoproteins in the mediation of epithelial invasion during pneumococcal colonisation. Targeting mucosal innate immunity and epithelial microinvasion alongside the induction of an adaptive immune response may be effective in preventing pneumococcal colonisation and disease.

microbiology↗

bioRxiv2023

Essential role of proline synthesis and the one-carbon metabolism pathways for systemic virulence of Streptococcus pneumoniae

Previous virulence screens have indicated potential roles during Streptococcus pneumoniae infection for the one-carbon metabolism pathway component Fhs and proline synthesis mediated by ProABC. To define how these metabolic pathways affect S. pneumoniae virulence we have investigated phenotypes and transcription profiles of {Delta}fhs and {Delta}proABC strain mutants. S. pneumoniae capsular serotype 6B BHN418 {Delta}fhs and {Delta}proABC mutant strains were markedly reduced virulence in mouse models of systemic infection and pneumonia, but were still able to colonise the nasopharynx. Although the {Delta}fhs and {Delta}proABC mutant strains grew normally in complete media, both mutant strains had markedly impaired growth in chemically defined medium, human serum and human CSF. The {Delta}proABC strain also had impaired growth under conditions of osmotic and oxidative stress. When transferred to the serotype 2 D39 S. pneumoniae strain background, the {Delta}fhs mutation replicated the virulence and growth in serum phenotype of the BHN418 mutation. In contrast, the D39 {Delta}proABC mutant could cause septicaemia and grow in human serum, indicating the role of this genetic locus during virulence is strain-specific. In human sera the {Delta}fhs and {Delta}proABC mutants both had major derangements in global gene transcription affecting multiple but different metabolic pathways, identifying the corresponding S. pneumoniae metabolic functions affected by these genes under infection-related conditions. Our data demonstrate an essential role for the S. pneumoniae one- carbon metabolism and a strain-conditional role for proline biosynthesis for growth in physiological fluids and therefore systemic infection, and further demonstrate the vital importance of bacterial metabolism for disease pathogenesis. ImportanceRapid adaptation to grow within the physiological conditions found in the host environment is an essential but poorly understood virulence requirement for systemic pathogens such as Streptococcus pneumoniae. We have now demonstrated an essential role for the one-carbon metabolism pathway and a conditional role depending on strain background for proline biosynthesis for S. pneumoniae growth in serum or CSF and therefore for systemic virulence. RNAseq data demonstrated that loss of one carbon metabolism or proline biosynthesis both have profound but differing effects on S. pneumoniae metabolism in human serum, identifying the metabolic processes dependent on each pathway during systemic infection. These data provide a more detailed understanding of the adaptations required by systemic bacterial pathogens in order to cause infection, and demonstrate that the requirement for some of these adaptations vary between strains from the same species and could therefore underpin strain variations in virulence potential.

microbiology↗